Project Summary/Abstract: Much of current molecular oncology research is grounded on the paradigm that the behavior of cancer cells and the tumors that they form is dictated by the mutant genomes that these cells have acquired en route to entering into neoplastic/cancerous states. Implicit in this work is the notion that non-genetic mechanisms (that are not directly influenced by DNA sequence) ? notably changes organized by non-genetic (i.e., epigenetic) regulatory circuits operating within a cancer cell ? play a secondary role in dictating its behavior. In fact, the majority of cancer cell traits may actually be governed by non-genetic regulatory programs that operate in a relatively stable fashion to ensure that many of these traits are transmitted heritably from normal cells-of-origin through multiple cancer cell generations during the course of multi-step tumor development. In addition, activation within carcinoma cells of the cell-biologic program termed the epithelial-mesenchymal transition (EMT) creates a second dimension of cancer cell behavior that is not dictated by the cells' DNA sequences, acting by converting relatively benign carcinoma cells to malignant derivatives. These two non- genetic regulatory mechanisms exert profound effects on the malignant progression of cancer cells and their responsiveness to various forms of therapy. However, relatively little is known about how these non- genetically determined traits of cancer cells are acquired and disrupted during tumor formation. Super-enhancers (SEs) represent large aggregations of transcription factors (TFs) associated with a relatively small number (several hundred) of gene promoters in both normal cells and cancer cells. These SEs are responsible for orchestrating the differentiation programs of a variety of normal cell types that enables them to become tissue-specialized cell types. The proposed research examines the disruptive effects of specific oncogene-encoded proteins (and/or loss of tumor suppressor gene proteins) on the spectrum of SEs within experimentally transformed human cells from a variety of normal cell lineages, with the goal of understanding which SEs survive disruption and continue to influence the behavior of resulting tumorigenic cells (with respect to their responsiveness to therapy and their malignant traits), and which are altered, resulting in the acquisition of novel cancer cell traits. As a complement to these analyses will be an examination of the SEs associated with the EMT programs activated in various types of transformed cells, these being activated either through the induced expression of EMT-inducing TFs or the exposure of transformed cells to known EMT-inducing growth factors and cytokines. By surveying SEs in normal cell types and their neoplastic derivatives, the proposed work will generate an experimental platform that will enable many researchers to finally elucidate at the molecular level why and how various distinct subtypes of human carcinomas naturally exhibit an array of key biological traits or do so in response to imposed cytotoxic therapies.